Summary: We established that oligodeoxynucleotides expressing "CpG" motifs (CpG ODN)mediate a variety of immunological functions, including the activation of B lymphocytes and the induction of Th1 cytokine production by NK, T and dendritic cells. We recently established that "D" type ODN could support the maturation of human peripheral blood monocytes into antigen presenting dendritic cells. We have been exploring the mechanism(s) underlying this immune activation, and found that a 6 base pair DNA motif (containing an unmethylated CpG dinucleotide flanked by two 5' purines and two 3' pyrimidines) that is common to bacterial but not mammalian DNA caused this stimulation in mice. In humans, we found that different motifs (and different types of nucleotide backbones) were required to induce immune activation. We've identified two broad and distinct categories of ODN ("D" and "K" type) that induce different immune responses on human PBMC in vitro and in rhesus macagues in vivo. This induction of different types of immune response may prove useful in optimizing the human response specific pathogens. We found that synthetic oligonucleotides expressing CpG motifs can act as anti-allergens (by deviating the immune response towards Th1 and away from Th2 driven IgE secretion), and can be used to prevent and/or treat bacterial, parasitic and viral infections by up-regulating the innate immune system. We are therefore examining methods of increasing and prolonging the protective effects of CpG ODN. Most recently, we demonstrated that CpG ODN were able to provide some protection against infection by anthrax and Ebola virus in murine models. We also showed that we could significantly reduce the level of Leishmania infection in rhesus monkeys treated with CpG ODN expected to be active in humans. By analyzing the response of >100 normal donors to a large panel of ODN, we found that individuals varied in their response to specific ODN, such that optimal stimulation of a diverse population will require combinations of different ODN motifs. We've also shown that PBMC from patients infected with HIV continue to benefit from the stimulatory activity of CpG ODN. A series of studies identified those regions of the IL-6 and IL-12 promoter that are involved in CpG ODN induced cell triggering at the molecular level. We also identified Toll-like receptor 9 as being critical for cellular recognition of "K" type CpG motifs by human cells, and accumulated data suggesting that cellular activation may proceed through protein kinase C. We've also been exploring the role of "suppressive motifs" present in mammalian DNA, and their ability to regulate the innate immune response elicited by CpG motifs. These suppressive motifs may play a critical role insuring that over-exhuberant responses to CpG ODN do not harm the host.